Degradable macromolecular magnetic resonance imaging contrast agents and methods thereof

ABSTRACT

Novel degradable macromolecular magnetic resonance imaging contrast agents for use in various diagnostic procedures, and methods for synthesizing, using and degrading these agents, are disclosed. The macromolecule contrast agents disclosed in various aspects of this invention are degradable gadolinium compounds which show prolonged plasma retention, and enhanced permeability and retention in solid tumors, but are still capable of being rapidly cleared from the body.

CROSS-REFERENCE TO RELATED APPLICATIONS

Applicants claim priority under 35 U.S.C. § 119 from U.S. ProvisionalPatent Application No. 60/338,831, filed Nov. 13, 2001, and entitled“Degradable Macromolecular Magnetic Resonance Imaging Contrast Agents,And Methods Thereof.”

BACKGROUND OF THE INVENTION

1. Field of the Invention

This invention relates generally to macromolecular contrast agents usedin diagnostic imaging, and methods of synthesizing, using and degradingsuch compounds.

2. Description of the Related Art

Magnetic resonance imaging (MRI) is a non-invasive method for medicaldiagnosis. Paramagnetic metal complexes are often used as contrastagents to enhance the image contrast between normal tissue and diseasedtissue. Paramagnetic metal ions that are typically used in diagnosticprocedures include manganese (Mn²⁺), iron (Fe⁺³), and gadolinium (Gd⁺³).Chelates of Gd⁺³ are frequently used as MRI contrast agents because oftheir long electronic relaxation time and high magnetic moment.Gadolinium-based contrast agents include small molecular gadoliniumcomplexes such as Gd(III)(DTPA) [diethylenetriaminepentaacetate] andGd(III)(DOTA) [1,4,7,10-tetraazadodecanetetraacetic acid], and theirderivatives. Use of stable, non-toxic paramagnetic chelates inconjunction with various diagnostic procedures, such as MRI, have beenused in the prior art to increase the accuracy of diagnosis.

Macromolecular MRI contrast agents, sometimes referred to aslong-circulating or blood pool agents, are particularly useful becauseof their prolonged retention time in the blood pool. These agentstypically increase the magnetic resonance signal of blood by shorteningthe T₁ relaxation time of blood and can be used for a variety ofdiagnostic procedures, including magnetic resonance angiography. Becauseof the enhanced permeability and retention, these contrast agentsaccumulate effectively in solid tumors and have a potential in contrastenhancement in MR cancer imaging. However, the macromolecular contrastagents currently available to the medical community have poor clearancerates. As a result of poor clearance rates, use of Gd macromolecularagents may result in Gd accumulation in bone and other tissues,resulting in toxicity and adverse side-effects.

SUMMARY OF THE INVENTION

It is an object of several embodiments of the present invention toprovide novel degradable macromolecular contrast agents which have thecharacteristics of prolonged blood circulation time and enhancedpermeability and retention in solid tumors, but are still capable ofbeing rapidly cleared from the body after diagnostic exams. In apreferred embodiment, these contrast agents are gadolinium complexesthat are used in magnetic resonance imaging. In an alternativeembodiment, the macromolecular Gd complexes are used in X-ray computedtomography. In other embodiments, the degradable polymer ligands canform chelates with radioactive metal ions for scintigraphy, positronemission tomography and radiotherapy.

In another aspect of the invention, the complexes include targetingmolecules. The incorporation of targeting molecules, including, but notlimited to, antibodies, antibody fragments, peptides, and other proteinsresults in macromolecular contrast agents with targeting ability.

It is another object of various aspects of the invention to provide amethod for obtaining a magnetic resonance image of a tissue or organ ofa mammal by administering one or more macromolecule contrast agents andobtaining a magnetic resonance image. In several embodiments, themacromolecule contrast agents are capable of being degraded by bothendogenous and exogenous compounds. In one embodiment, the contrastagents are degraded by endogenous mercaptans.

It is a further object of various embodiments to degrade or stimulatedegradation of the macromolecule contrast agents by administering one ormore disulfide bond reducing compounds or other compounds that stimulatethe degradation of the macromolecule contrast agents. In one embodiment,exogenous mercaptans are delivered to the mammal. Another object ofseveral embodiments of the current invention is to administermacromolecule contrast agents in conjunction with other agents. Invarious aspects, physiologically acceptable agents, such as diluents andcarriers, are also administered.

In a preferred embodiment, the current invention is a macromoleculecontrast formulation comprising any one or more of the followingmacromolecule contrast agents, or physiologically acceptable saltsthereof:

-   -   wherein R and R′ are independently selected from the group        consisting of C₁ to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X and Y are independently selected from the group        consisting of O and NH; and    -   wherein n is an integer between 2 and 10,000;    -   wherein R and R′ are independently selected from the group        consisting of C₁ to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X and Y are independently selected from the group        consisting of amide groups, ester groups, urea groups, thiourea        groups, carbonate groups, carbamate groups, ether bonds and        thioether bonds;    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, polysaccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates;    -   wherein n is an integer between 2 and 10,000; and    -   wherein L is selected from the group consisting of        diethylenetriaminepentaacetate (DTPA) or its derivatives,        1,4,7,10-tetraazadodecanetetraacetate (DOTA) and its        derivatives, 1,4,7,10-tetraazadodecane-1,4,7-triacetate (DO3A)        and its derivatives, ethylenediaminetetraacetate (EDTA) and its        derivatives, 1,4,7,10-tetraazacyclotridecanetetraacetic acid        (TRITA) and its derivatives,        1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid        (TETA) and its derivatives,        1,4,7,10-tetraazadodecanetetramethylacetate (DOTMA) and its        derivatives, 1,4,7,10-tetraazadodecane-1,4,7-trimethylacetate        (DO3MA) and its derivatives,        N,N′,N″,N′″-tetraphosphonatomethyl-1,4,7,10-tetraazacyclododecane        (DOTP) and its derivatives,        1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(methylene        methylphosphonic acid) (DOTMP) and its derivatives,        1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(methylene        phenylphosphonic acid) (DOTPP) and its derivatives and other        chelating ligands;    -   wherein R and R′ are independently selected from the group        consisting of C, to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X is selected from the group consisting of amide groups,        ester groups, and ether bonds;    -   wherein Y is selected from the group consisting of O and NH;    -   wherein n is an integer between 2 and 10,000; and    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, polysaccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates;    -   wherein R and R′ are independently selected from the group        consisting of C₁ to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X is selected from the group consisting of amide groups,        ester groups, and ether bonds;    -   wherein Y is selected from the group consisting of O and NH;    -   wherein n is an integer between 2 and 10,000; and    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, saccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates;    -   wherein R and R′ are independently selected from the group        consisting of C, to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X is selected from the group consisting of amide groups,        ester groups, and ether bonds;    -   wherein Y is selected from the group consisting of O and NH;    -   wherein n is an integer between 2 and 10,000; and    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, polysaccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates;    -   wherein R, R′, R″ and R′″ are independently selected from the        group consisting of H, C₁ to C₁₈ alkyls, substituted alkyls,        aryls, one or more functional groups containing alkyl, aryl or        polyethylene glycol, saccharides, amino acids, peptides,        proteins, peptide conjugates and protein conjugates;    -   wherein X is selected from the group consisting of O and NH; and    -   wherein n is an integer between 2 and 10,000;    -   wherein R and R′ are independently selected from the group        consisting of H, C₁ to C₁₈ alkyls, substituted alkyls, aryls,        one or more functional groups containing alkyl, aryl or        polyethylene glycol, saccharides, amino acids, peptides,        proteins, peptide conjugates and protein conjugates;    -   wherein X is selected from the group consisting of O and NH; and    -   wherein n is an integer between 2 and 10,000.

One embodiment comprises one or more of the following molecular contrastagents:

-   -   wherein n is an integer between 2 and 10,000; and    -   wherein x is an integer between 1 and 20; and    -   wherein m is an integer between 0 and 10,000.

One aspect of the invention includes a method for obtaining a magneticresonance image of a tissue or organ of a mammal by administering aneffective amount of one or more macromolecule contrast agents to themammal and obtaining a magnetic resonance image. In a preferredembodiment, one or more of the macromolecule contrast agents aredegraded by endogenous mercaptans.

According to one aspect of the invention, one or more compounds thatstimulate the degradation of said macromolecule contrast agent is alsoadministered. In another aspect, one or more disulfide bond reducingcompounds is also administered. The disulfide bond reducing compound isselected from the group consisting of one or more of the following:mercaptans, NADH, NADPH, hydrazines, phosphines, zinc, tin(II), sodiumsulfide, performic acid, hydrogen peroxide.

Mercaptans used in various aspects of the present invention are selectedfrom the group consisting of one or more of the following: cysteine andits derivatives, glutathione and its derivatives, cysteinylglycine andits derivatives, 2,3-dimercaptosuccinic acid and its derivatives,2,3-dimercapto-1-propanesulfonic acid and its derivatives,2-mercaptoethanol, penicillamine and its derivatives, mercaptoaceticacid and its derivatives, mercaptoanisole, 2-mercaptobenzoic acid andits derivatives, 4-mercaptobenzoic acid and its derivatives,2-mercapto-5-benzimidazolesulfonic acid and its derivatives,2-mercaptobenzothiazole, 3-mercapto-iso-butyric acid,mercaptocyclohexane, 2-mercaptoethanesulfonic acid,2-mercaptoethylamine, 2-mercaptoethylamine hydrochloride,3-mercapto-1,2-propanediol, 3-mercapto-1-propanesulfonic acid,3-mercapto-1-propanol, 2-mercaptopropionic acid, 3-mercaptopropionicacid, diethyldithiocarbamate, dithioerythritol, and dithioglycol.

According to one aspect of the invention, two or more of macromoleculecontrast agents are administered simultaneously. In one embodiment, themacromolecule contrast formulation comprises a first macromoleculecontrast agent and a second macromolecule contrast agent, wherein thesecond macromolecule contrast agent is administered after theadministration of said first macromolecule contrast agent.

In another embodiment, at least one of the macromolecule contrast agentsis administered in conjunction with one or more physiologicallyacceptable agents selected from the group consisting of: diluents,carriers, antibodies, antibody Fab′ fragments, antibody F(ab′)₂fragments, and delivery systems.

In a preferred embodiment, at least one of the macromolecule contrastagents is administered in conjunction with one or more contrast agentsselected from the group consisting of: paramagnetic metal complexes,radioactive metal complexes, therapeutic agents, proteins, DNA, RNA,drug delivery systems and gene delivery systems.

It is an object of several embodiments of the current invention toobtain a magnetic resonance image of healthy or tumorous tissues ororgans, including but not limited to, liver, spleen, lung, heart,kidney, tumors, ovary, pancreas, biliary system, peritoneum, muscles,head, neck, esophagus, bone marrow, lymph node, lymph vessels, nervoussystem, brain, spinal cord, blood capillaries, stomach, small intestine,and large intestine.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows size exclusion chromatograms of DPTA-cystamine copolymer,(DPTA-Gd)-cystamine copolymer and the degradation mixtures of(DPTA-Gd)-cysteine copolymer with cysteine at 37° C. for 10 minutes and30 minutes, respectively.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

In a preferred embodiment of the present invention, degradablemacromolecular MRI contrast agents having degradable disulfide bond inthe polymer backbone are disclosed. An alternative embodiment includesdegradable macromolecular MRI contrast agents which contain a structurethat conjugates the paramagnetic metal complexes, Gd(DTPA), Gd(DOTA) ortheir derivatives, to polymer or protein carriers through a disulfidebond. In several embodiments, the disulfide bond can readily be reducedby one or more mercaptans or thiols, including, but not limited tocysteine, N-acetylcysteine, glutathione, and the like. These mercaptansmay be endogenously available or exogenously delivered. Alternatively,compounds that stimulate the endogenous degradation of macromoleculecontrast agents may also be delivered using several embodiments of thecurrent invention, including, but not limited to, cysteine,N-acetylcysteine, glutathione, cysteinylglycine, and homocysteine. Oneskilled in the art will appreciate that several agents capable ofreducing disulfide bonds may be used according to various aspects ofthis invention, including, but not limited to cysteine,N-acetylcysteine, glutathione, cysteinylglycine, and their derivatives.

Another object of several embodiments is to provide a method forobtaining a magnetic resonance image of a tissue or organ of a mammal byadministering an effective amount of one or more macromolecule contrastagents to the mammal and obtaining a magnetic resonance image. Oneskilled in the art will understand that many known methods of obtainingmagnetic resonance image exist in the scientific and medical field. In apreferred embodiment, an MRI procedure is performed on a human subject.One skilled in the art will appreciate that a variety of tissues andorgans may be examined using different aspects of this invention,including, but not limited to, liver, spleen, lung, esophagus, bonemarrow, lymph node, lymph vessels, nervous system, brain, spinal cord,blood capillaries, stomach, small intestine, large intestine. Oneskilled in the art will appreciate that both normal tissues and abnormaltissues, such as tumors, can be examined.

Another aspect of this invention relates to a method of clearing metalcomplexes. Preferably, the clearance procedure is performed after theMRI procedure has been completed or substantially completed. In oneembodiment, mercaptans, or other similar agents, are administered afterthe MRI procedure. In various embodiments, these agents facilitate theclearing process by cleaving the macromolecular backbone. Alternatively,or in addition, clearing occurs by removal of the paramagnetic metalcomplexes from the polymer carriers by cleavage of the disulfide bond.Several embodiments are particularly advantageous because theparamagnetic metal complexes released from the macromolecules can becleared at a rate comparable to that of the small molecular contrastagents used clinically today.

In a preferred embodiment, the macromolecular compounds have a prolongedretention time in the blood pool, favorable accumulation in the solidtumor tissues, and are cleared rapidly after MRI. These macromolecularagents, and the methods described thereof, will be indispensable toolsin a variety of medical procedures, including, but not limited to,angiography, plethysmography, lymphography, mammography, cancerdiagnosis, and functional and dynamic MRI.

Chemical Compounds

The macromolecular MRI contrast agents disclosed in accordance withvarious embodiments of the present invention are represented by thefollowing generic formulae:

-   -   wherein R and R′ are independently selected from the group        consisting of C₁ to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X and Y are independently selected from the group        consisting of O and NH; and    -   wherein n is an integer between 2 and 10,000.    -   wherein R and R′ are independently selected from the group        consisting of C₁ to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X and Y are independently selected from the group        consisting of amide groups, ester groups, urea groups, thiourea        groups, carbonate groups, carbamate groups, ether bonds and        thioether bonds;    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, polysaccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates;    -   wherein n is an integer between 2 and 10,000; and    -   wherein L is selected from the group consisting of        diethylenetriaminepentaacetate (DTPA) or its derivatives,        1,4,7,10-tetraazadodecanetetra-acetate (DOTA) and its        derivatives, 1,4,7,10-tetraazadodecane-1,4,7-triacetate (DO3A)        and its derivatives, and any other chelating ligands.    -   wherein R and R′ are independently selected from the group        consisting of C, to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X is selected from the group consisting of amide groups,        ester groups, urea groups, thiourea groups, carbonate groups,        carbamate groups, ether bonds and thioether bonds;    -   wherein Y is selected from the group consisting of O and NH;    -   wherein n is an integer between 2 and 10,000; and    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, polysaccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates.    -   wherein R and R′ are independently selected from the group        consisting of C₁ to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X is selected from the group consisting of amide groups,        ester groups, urea groups, thiourea groups, carbonate groups,        carbamate groups, ether bonds and thioether bonds;    -   wherein Y is selected from the group consisting of O and NH;    -   wherein n is an integer between 2 and 10,000; and    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, polysaccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates.    -   wherein R and R′ are independently selected from the group        consisting of C₁ to C₁₈ alkyls, substituted alkyls, aryls, one        or more functional groups containing alkyl, aryl or polyethylene        glycol, saccharides, amino acids, peptides, proteins, peptide        conjugates and protein conjugates;    -   wherein X is selected from the group consisting of amide groups,        ester groups, urea groups, thiourea groups, carbonate groups,        carbamate groups, ether bonds and thioether bonds;    -   wherein n is an integer between 2 and 10,000; and    -   wherein P is selected from the group consisting of water soluble        polymer chains, dendrimers, polysaccharides, peptides, proteins,        polymer-peptide conjugates and polymer-protein conjugates.    -   wherein R, R′, R″ and R′″ are independently selected from the        group consisting of H, C₁ to C₁₈ alkyls, substituted alkyls,        aryls, one or more functional groups containing alkyl, aryl or        polyethylene glycol, saccharides, amino acids, peptides,        proteins, peptide conjugates and protein conjugates;    -   wherein X is selected from the group consisting of O and NH; and    -   wherein n is an integer between 2 and 10,000.    -   wherein R and R′ are independently selected from the group        consisting of H, C₁ to C₁₈ alkyls, substituted alkyls, aryls,        one or more functional groups containing alkyl, aryl or        polyethylene glycol, saccharides, amino acids, peptides,        proteins, peptide conjugates and protein conjugates;    -   wherein X is selected from the group consisting of O and NH; and    -   wherein n is an integer between 2 and 10,000.

The macromolecular MRI contrast agents disclosed in accordance withvarious embodiments of the present invention are represented by thefollowing formulae:

-   -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000.    -   wherein n is an integer between 2 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein m is an integer between 0 and 10,000.    -   wherein n is an integer between 2 and 10,000; and    -   wherein x is an integer between 1 and 20; and    -   wherein m is an integer between 0 and 10,000.        Synthesis Reactions        Synthesis of (DTPA-Gd)-cystamine Copolymer

Cystamine dichloride (0.63 g, 2.8 mmol) and triethylamine (2 ml, excess)were mixed with 20 ml DMSO (anhydrous), followed by DTPA dianhydride (1g, 2.8 mmol). The mixture was stirred at room temperature for 2 days.The product was precipitated in acetone. The precipitate was collectedby filtration and washed with acetone. The product was then dissolved indeionized water and dialyzed (MWCO 6,000–8,000 Da) for 2 days. Thesolution was then lyophilized, giving 550 mg (34%) DTPA-cystaminecopolymer, indicated as (A) below. The copolymer was characterized bysize exclusion chromatography (FIG. 1). DTPA-cystamine copolymer (300mg) was dissolved in 2 ml DI water and GdCl₃ was added with xyleneorange as an indicator. The solution was neutralized with NaOH to pH5.0. The resulting solution was passed through a Sephadex G-25 column toremove excess GdCl₃ and salt. Finally, the product was lyophilizedgiving a colorless product, (DTPA-Gd) cystamine copolymer, indicated as(B) below. The yield was 250 mg, or 83%. The product was characterizedby size exclusion chromatography (SEC), FIG. 1.

Synthesis of HPMA Copolymer-(DTPA-Gd) Conjugate

Cystamine hydrochloride (400 mg excess) and triethylamine (1.0 ml,excess) were dissolved in 20 ml methanol, and HPMA copolymer withp-nitrophenyl active ester (Mn=18,300 Da and Mw=22,500 Da, ONp %=9.6mol-%) (420 mg) in 5 ml methanol was added dropwise to above solutionwith stirring. The mixture was stirred overnight at room temperature.The solution was concentrated under vacuum and applied to Sephadex LH-20column to separate the polymer conjugate, eluted with methanol. Thepolymer fraction was collected and solvent was removed under vacuumgiving HPMA copolymer-cystamine conjugates. The product was dissolved inwater, and DTPA dianhydride (300 mg, excess) was added to the solution.The mixture was stirred for 30 minutes, and pH was adjusted to 7.0 withNaHCO₃. The mixture was stirred at room temperature overnight. Thepolymer was separated from excess DTPA by passing through a SephadexG-25 column, eluted with water. GdCl₃ was added with xylene orange as anindicator. The solution was neutralized with NaOH to pH 5.0. Theresulting solution was passed through a Sephadex G-25 column to removeexcess GdCl₃ and salt. Finally, the product was lyophilized giving acolorless product, HPMA copolymer-[DTPA-Gd] conjugate, indicated as (C)below. The yield was 340 mg, or 85%.

Degradation of (DTPA-Gd)-cystamine Copolymer

(DTPA-Gd)-cystamine copolymer (5 mg) was dissolved in PBS buffer (1.0ml, pH 7.4) and incubated with cysteine (1 mM) at 37° C. for 10 min and30 min, respectively. The degradation reaction, (D), is shown below. Thereaction mixture was analyzed by size exclusion chromatography with ananalytical Superose 6 column, FIG. 1. Results indicated that themacromolecular MRI contrast agent, (DTPA-Gd)-cystamine copolymer, wascompletely degraded into small molecules by cysteine during a very shortincubation period.

Suitable dosages of the macromolecule contrast agents disclosed inseveral embodiments of the current invention will generally range fromabout 0.001 mmol to about 10 mmol gadolinium per kg body weight, morepreferably, between about 0.01 mmol and 1.0 mmol gadolinium per kg bodyweight. The compounds are preferably administered systemically, asappropriately determined by one skilled in the art. More preferably, thecontrast agents are administered intravenously. However, these agentsmay be delivered subcutaneously, or by any other route which may beeffective in providing systemic dosing of the agent. In one embodimentof the current invention, two or more of the macromolecule contrastagents are administered simultaneously. In an alternative embodiment, asecond macromolecule contrast agent is administered after the deliveryof a first macromolecule contrast agent. In one embodiment of thepresent invention, one or more macromolecule contrast agents may beadministered in conjunction with one or more physiologically acceptablediluents, carriers or other contrast agents.

While a number of preferred embodiments of the invention and variationsthereof have been described in detail, other modifications and methodsof use will be readily apparent to those of skill in the art.Accordingly, it should be understood that various applications,modifications and substitutions may be made of equivalents withoutdeparting from the spirit of the invention or the scope of the claims.

1. A macromolecule contrast agent comprising a compound according to thefollowing formula:

wherein R and R′ are independently selected from the group consisting ofC₁ to C₁₈ alkyls, substituted alkyls, aryls, polyethylene glycol,saccharides, amino acids, peptides, proteins, peptide conjugates andprotein conjugates, and combinations of alkyl, aryl and polyethyleneglycol; wherein X and Y are independently selected from the groupconsisting of O and NH; and wherein n is an integer between 2 and10,000.
 2. The macromolecule contrast agent of claim 1, wherein theagent has the following formula:

wherein R, R′, R″ and R′″ are independently selected from the groupconsisting of H, C₁ to C₁₈ alkyls, substituted alkyls, aryls,polyethylene glycol, saccharides, amino acids, peptides, proteins,peptide conjugates and protein conjugates, and combinations of alkyl,aryl and polyethylene glycol; wherein X is selected from the groupconsisting of O and NH; and wherein n is an integer between 2 and10,000.
 3. The macromolecule contrast agent of claim 1, wherein theagent has the following formula:

wherein R and R′ are independently selected from the group consisting ofH, C₁ to C₁₈ alkyls, substituted alkyls, aryls, polyethylene glycol,saccharides, amino acids, peptides, proteins, peptide conjugates andprotein conjugates, and combinations of alkyl, aryl, and polyethyleneglycol; wherein X is selected from the group consisting of O and NH; andwherein n is an integer between 2 and 10,000.
 4. A macromoleculecontrast agent comprising a compound selected from the group consistingof compounds according to the following formulae:


5. The macromolecule contrast agent of claim 1, wherein the agent hasthe following structure:

wherein n is an integer between 2 and 10,000.
 6. The macromoleculecontrast agent of claim 1, wherein the agent has the followingstructure:

wherein n is an integer between 2 and 10,000.